Mammary Carcinogenesis: The Role of Pleiotrophin. Breast cancer growth and metastasis requires growth factor signals between the tumor cells and the normal surrounding host tissue. We propose that the secreted polypeptide growth factor pleiotrophin (PTN) plays a major role in mammary carcinogenesis as well as in the growth and metastasis of breast cancer. This hypothesis is based on the biological effects of PTN expression in selected tumors models, the high levels of PTN mRNA in 60% of tumor samples from breast cancer patients and on its upregulation during carcinogen-induced rodent mammary cancer. Furthermore, PTN is highly expressed in immature mouse mammary glands that are sensitive to carcinogens and is downregulated permanently in mice that have undergone pregnancy and are protected against carcinogenesis. Finally, the activity of PTN on endothelial cells in vitro suggests that PTN can serve as an angiogenesis factor during tumor progression. In support of this role of PTN as an angiogenesis factor, reduction of constitutively expressed endogenous rib mRNA in human tumor cell lines by PTN-targeted ribozymes reduced tumor growth and metastasis even when in vitro growth of the tumor cells remained unaffected. Based on these preliminary studies, we wish to elucidate the role of PTN in breast cancer progression and in mammary carcinogenesis with the ultimate goal to develop better prognostic markers as well as novel therapeutic strategies. Under aim 1 we will study whether serum levels of PTN in breast cancer patients can predict long term outcome of their disease an-or the response to therapy. We will use samples from patients with over 10 years of follow-up from our serum bank core facility for this analysis. Under aim 2 we will study to what extent hormonal regulation affects PTN expression in mammary gland maturation and during mammary carcinogenesis in rodent models. We will study the influence of steroid hormones and retinoid acid receptor ligands on PTN expression and on mammary gland biology or carcinogenesis in animals and in organ culture. Under aim 3 we will study to what extent sustained high levels of expression of rib will affect mammary gland maturation, hormonal responsiveness and susceptibility to carcinogens at different stages of mammary gland maturation. To achieve this, we will express PTN as a transgene in mice under a tetracycline-regulated promoter. We will study the effects of a challenge of the transgenic animals (and of organ explants) with hormonal stimuli as well as with carcinogens. Under aim 4 we will study to what extent reduction of endogenous PTN expression will affect mammary gland physiology and pathology. We will use ribozyme-targeting of rib to achieve this goal. We will generate mice expressing PTN-targeted ribozymes under the control of a tetracycline-regulated promoter and study spontaneous gland maturation during pregnancy as well as the effects of hormonal and carcinogen challenges with the endogenous PTN suppressed by the ribozyme transgene. Under aim 5 we will develop gene transfer vectors to target rib expression with ribozymes in vivo. We will use adenovirus- and herpes virus-based vectors for gene transfer.